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Context-dependent changes in genetic interactions are an important feature of cellular pathways and their varying responses under different environmental conditions. However, methodological frameworks to investigate the plasticity of genetic interaction networks over time or in response to external stresses are largely lacking. To analyze the plasticity of genetic interactions, we performed a combinatorial RNAi screen in Drosophila cells at multiple time points and after pharmacological inhibition of Ras signaling activity. Using an image-based morphology assay to capture a broad range of phenotypes, we assessed the effect of 12768 pairwise RNAi perturbations in six different conditions. We found that genetic interactions form in different trajectories and developed an algorithm, termed MODIFI, to analyze how genetic interactions rewire over time. Using this framework, we identified more statistically significant interactions compared to end-point assays and further observed several examples of context-dependent crosstalk between signaling pathways such as an interaction between Ras and Rel which is dependent on MEK activity. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter ). Within a cell, communication routes that involve many different genes work to control how the cell responds to the environment. Although different communication routes – so called signaling pathways – control different cell activities, they do not work in isolation. Instead, they form part of larger regulatory networks that maintain the cell in an appropriate state. As such, changing the activity of one pathway may in turn affect another seemingly unrelated pathway. The Ras signaling pathway helps to control when cells divide. When this signaling is not regulated correctly, cells can start to divide uncontrollably, leading to cancer. Drugs that suppress the activity of overactive Ras pathways could help to treat cancer. But how do the wider regulatory networks in the cell rewire themselves over time in response to this treatment? To investigate this question, Heigwer et al. used a method called RNA interference to alter the activity of different pairs of 168 genes in fruit fly cells that had been grown in the laboratory. This meant 12,768 gene interactions were examined in total. Some of the cells had been treated with a drug that suppresses Ras signaling. By developing a new cell imaging and analysis system, Heigwer et al. could examine how the cell’s regulatory networks were affected by the drug at three different time points after treatment. The results show that housekeeping genes, which handle basic cell duties, take more time to rewire their interactions than signaling pathways. Heigwer et al. also developed a computational method – called MODIFI – to analyze how environment and time affect how genes interact. This highlighted a number of signaling pathways that are strongly affected by the suppression of Ras signaling, including an unexpected immune signaling pathway. In the future, more research will be needed to study the context-dependency of interactions between genetic networks in different cell types and in living organisms. A better understanding of this context-dependency will be important to understand how cancerous cells develop drug resistance. The data collected by Heigwer et al. could also be used by other researchers to explain any unexpected gene interactions that affect the signaling pathways they are studying.

Characterization of spatial and temporal variation in grassland productivity and nutrition is crucial for a comprehensive understanding of ecosystem function. Although within-site heterogeneity in soil and plant properties has been shown to be relevant for plant community stability, spatiotemporal variability in these factors is still understudied in temperate grasslands. Our study aimed to detect if soil characteristics and plant diversity could explain observed small-scale spatial and temporal variability in grassland productivity, biomass nutrient concentrations, and nutrient limitation. Therefore, we sampled 360 plots of 20 cm × 20 cm each at six consecutive dates in an unfertilized grassland in Southern Germany. Nutrient limitation was estimated using nutrient ratios in plant biomass. Absolute values of, and spatial variability in, productivity, biomass nutrient concentrations, and nutrient limitation were strongly associated with sampling date. In April, spatial heterogeneity was high and most plots showed phosphorous deficiency, while later in the season nitrogen was the major limiting nutrient. Additionally, a small significant positive association between plant diversity and biomass phosphorus concentrations was observed, but should be tested in more detail. We discuss how low biological activity e.g., of soil microbial organisms might have influenced observed heterogeneity of plant nutrition in early spring in combination with reduced active acquisition of soil resources by plants. These early-season conditions are particularly relevant for future studies as they differ substantially from more thoroughly studied later season conditions. Our study underlines the importance of considering small spatial scales and temporal variability to better elucidate mechanisms of ecosystem functioning and plant community assembly.

Phylogenetic diversity (PD) has been successfully used as a complement to classical measures of biological diversity such as species richness or functional diversity. By considering the phylogenetic history of species, PD broadly summarizes the trait space within a community. This covers amongst others complex physiological or biochemical traits that are often not considered in estimates of functional diversity, but may be important for the understanding of community assembly and the relationship between diversity and ecosystem functions. In this study we analyzed the relationship between PD of plant communities and land-use intensification in 150 local grassland plots in three regions in Germany. Specifically we asked whether PD decreases with land-use intensification and if so, whether the relationship is robust across different regions. Overall, we found that species richness decreased along land-use gradients the results however differed for common and rare species assemblages. PD only weakly decreased with increasing land-use intensity. The strength of the relationship thereby varied among regions and PD metrics used. From our results we suggest that there is no general relationship between PD and land-use intensification probably due to lack of phylogenetic conservatism in land-use sensitive traits. Nevertheless, we suggest that depending on specific regional idiosyncrasies the consideration of PD as a complement to other measures of diversity can be useful.

OBJECTIVE To identify the source of a pseudo-outbreak of Mycobacterium gordonae DESIGN Outbreak investigation. SETTING University Hospital in Chicago, Ilinois. PATIENTS Hospital patients with M. gordonae-positive clinical cultures. METHODS An increase in isolation of M. gordonae from clinical cultures was noted immediately following the opening of a newly constructed hospital in January 2012. We reviewed medical records of patients with M. gordonae-positive cultures collected between January and December 2012 and cultured potable water specimens in new and old hospitals quantitatively for mycobacteria. RESULTS Of 30 patients with M. gordonae-positive clinical cultures, 25 (83.3%) were housed in the new hospital; of 35 positive specimens (sputum, bronchoalveolar lavage, gastric aspirate), 32 (91.4%) had potential for water contamination. M. gordonae was more common in water collected from the new vs. the old hospital [147 of 157 (93.6%) vs. 91 of 113 (80.5%), P=.001]. Median concentration of M. gordonae was higher in the samples from the new vs. the old hospital (208 vs. 48 colony-forming units (CFU)/mL; P<.001). Prevalence and concentration of M. gordonae were lower in water samples from ice and water dispensers [13 of 28 (46.4%) and 0 CFU/mL] compared with water samples from patient rooms and common areas [225 of 242 (93%) and 146 CFU/mL, P<.001]. CONCLUSIONS M. gordonae was common in potable water. The pseudo-outbreak of M. gordonae was likely due to increased concentrations of M. gordonae in the potable water supply of the new hospital. A silver ion-impregnated 0.5-μm filter may have been responsible for lower concentrations of M. gordonae identified in ice/water dispenser samples. Hospitals should anticipate that construction activities may amplify the presence of waterborne nontuberculous mycobacterial contaminants.

Distinguishing organic and conventional products is a major issue of food security and authenticity. Previous studies successfully used stable isotopes to separate organic and conventional products, but up to now, this approach was not tested for organic grassland hay and soil. Moreover, isotopic abundances could be a powerful tool to elucidate differences in ecosystem functioning and driving mechanisms of element cycling in organic and conventional management systems. Here, we studied the δ(15)N and δ(13)C isotopic composition of soil and hay samples of 21 organic and 34 conventional grasslands in two German regions. We also used Δδ(15)N (δ(15)N plant - δ(15)N soil) to characterize nitrogen dynamics. In order to detect temporal trends, isotopic abundances in organic grasslands were related to the time since certification. Furthermore, discriminant analysis was used to test whether the respective management type can be deduced from observed isotopic abundances. Isotopic analyses revealed no significant differences in δ(13)C in hay and δ(15)N in both soil and hay between management types, but showed that δ(13)C abundances were significantly lower in soil of organic compared to conventional grasslands. Δδ(15)N values implied that management types did not substantially differ in nitrogen cycling. Only δ(13)C in soil and hay showed significant negative relationships with the time since certification. Thus, our result suggest that organic grasslands suffered less from drought stress compared to conventional grasslands most likely due to a benefit of higher plant species richness, as previously shown by manipulative biodiversity experiments. Finally, it was possible to correctly classify about two third of the samples according to their management using isotopic abundances in soil and hay. However, as more than half of the organic samples were incorrectly classified, we infer that more research is needed to improve this approach before it can be efficiently used in practice.

Background In recent years, large-scale genetic screens using the CRISPR/Cas9 system have emerged as scalable approaches able to interrogate gene function with unprecedented efficiency and specificity in various biological contexts. By this means, functional dependencies on both the protein-coding and noncoding genome of numerous cell types in different organisms have been interrogated. However, screening designs vary greatly and criteria for optimal experimental implementation and library composition are still emerging. Given their broad utility in functionally annotating genomes, the application and interpretation of genome-scale CRISPR screens would greatly benefit from consistent and optimal design criteria. Results We report advantages of conducting viability screens in selected Cas9 single-cell clones in contrast to Cas9 bulk populations. We further systematically analyzed published CRISPR screens in human cells to identify single-guide (sg) RNAs with consistent high on-target and low off-target activity. Selected guides were collected in a novel genome-scale sgRNA library, which efficiently identifies core and context-dependent essential genes. Conclusion We show how empirically designed libraries in combination with an optimized experimental design increase the dynamic range in gene essentiality screens at reduced library coverage.

Both a high number of species and abundance in multiple trophic levels are required for ecosystems to continue to provide the services humans require of them. The importance of biodiversity in depth Numerous experiments have shown that the loss of biodiversity within single trophic groups — groups of organisms consuming resources from a similar level in the food chain — reduces the ability of ecosystems to deliver the services on which humans depend. How the loss of biodiversity in natural ecosystems consisting of multiple interacting trophic groups affects ecosystem functioning has remained unclear. Santiago Soliveres et al . have compiled data on the richness and abundance of 4,600 microbial, plant and animal taxa in 150 grasslands in Germany, together with information on 14 ecosystem services. Their analysis of the data demonstrates that biodiversity across multiple trophic groups is as important for the functioning of ecosystems as land-use intensity or environmental conditions. They conclude that the preservation of high levels of richness and diversity within a wide range of taxa will be key to ensuring that ecosystems continue to deliver the services on which humans rely. The findings also inform conservation and ecosystem management strategies by highlighting the most functionally relevant organisms, which include plants, soil bacteria and herbivorous insects. Many experiments have shown that loss of biodiversity reduces the capacity of ecosystems to provide the multiple services on which humans depend 1 , 2 . However, experiments necessarily simplify the complexity of natural ecosystems and will normally control for other important drivers of ecosystem functioning, such as the environment or land use. In addition, existing studies typically focus on the diversity of single trophic groups, neglecting the fact that biodiversity loss occurs across many taxa 3 , 4 and that the functional effects of any trophic group may depend on the abundance and diversity of others 5 , 6 . Here we report analysis of the relationships between the species richness and abundance of nine trophic groups, including 4,600 above- and below-ground taxa, and 14 ecosystem services and functions and with their simultaneous provision (or multifunctionality) in 150 grasslands. We show that high species richness in multiple trophic groups (multitrophic richness) had stronger positive effects on ecosystem services than richness in any individual trophic group; this includes plant species richness, the most widely used measure of biodiversity. On average, three trophic groups influenced each ecosystem service, with each trophic group influencing at least one service. Multitrophic richness was particularly beneficial for ‘regulating’ and ‘cultural’ services, and for multifunctionality, whereas a change in the total abundance of species or biomass in multiple trophic groups (the multitrophic abundance) positively affected supporting services. Multitrophic richness and abundance drove ecosystem functioning as strongly as abiotic conditions and land-use intensity, extending previous experimental results 7 , 8 to real-world ecosystems. Primary producers, herbivorous insects and microbial decomposers seem to be particularly important drivers of ecosystem functioning, as shown by the strong and frequent positive associations of their richness or abundance with multiple ecosystem services. Our results show that multitrophic richness and abundance support ecosystem functioning, and demonstrate that a focus on single groups has led to researchers to greatly underestimate the functional importance of biodiversity.

Introduction Cardiovascular disease (CVD) is the leading cause of death and the mortality rate and prognosis of CVD in women are worse compared to men. Adverse Pregnancy Outcomes (APOs) are frequently overlooked sex-specific risk factors for CVD and affect up to one in five pregnant women. This study evaluated healthcare providers'(HCPs) awareness of the long-term cardiovascular risk associated with gestational diabetes mellitus and hypertensive disorders of pregnancy. Methods A cross-sectional survey was conducted in Austria between March and August 2022 to assess HCPs’ knowledge, follow-up recommendations, and counseling regarding cardiovascular risk following APOs. The respondents were divided into general medicine, Obstetrics and Gynecology (O&G), general internal medicine, and cardiology. Results Of the 175 responses, 20% ( n  = 35) were from general medicine, 39% ( n  = 68) from O&G, 39% ( n  = 69) from general internal medicine and cardiology, and 2% ( n  = 3) from other specialties. Although most respondents were aware of increased CVD risk following APOs, significant knowledge gaps were identified, particularly concerning the prevalence and timing of CVD onset after APOs. Over 50% do not counsel women with APOs on cardiovascular risk reduction strategies and approximately half do not counsel on the risk of recurrence of APOs. Less than 20% provide women with written follow-up information. Differences in expertise were observed among specialties, with O&G demonstrating the highest level of knowledge. Conclusion This study identified knowledge gaps among HCPs in postpartum care for women with APOs highlighting the importance of standardized follow-up programs and the need for targeted education for HCPs. Graphical abstract

The population of adult CHD patients is continuously increasing. The underlying CHD affects performance and prognosis, but also has a significant impact on quality of life, psychosocial behaviour, anxiety and emotional disturbances. This study analyzes these parameters of patients after one or more heart operations and the possible psychological effects of medical and psychosocial complications at the Department of Cardiology of the Kepler University Hospital Linz. A total of 81 subjects participated in the questionnaire survey of the Institute of Cardiology and Clinical Psychology during their annual cardiological check-up. Of these, 80 participants were included in the study and three showed a mild CHD, 49 a moderate one, and 28 a severe one. This study has an exploratory design to assess possible stress factors and limitations in quality of life. For this purpose, a self-administered sociodemographic questionnaire and three standardised questionnaires were used. In summary, the quality of life of adult CHD is considered depending on the severity of the symptoms and compared with the healthy population. Differences in this regard are observed in individuals with lower symptom severity, who report higher psychological well-being. Sex differences are observed in physical role function and physical functioning. Based on the results, regular repetitions of the study, as well as continuous psychological and psychosocial support, are necessary, since challenges are predictable with the increasing age of adult CHD patients and since the upholding of good quality of life and dealing with difficult life circumstances must be supported.